Project Summary Although the properties of dissolved organic solutes and salts have been studied for over 130 years, we know little of the laws governing how they interact. Consider for example the fact that NaI can lead to an increase in the solubility of a protein (the Hofmeister Effect) or bring about a decrease in solubility and lead to precipitation of a protein (the Reverse Hofmeister Effect, RHE). This application concerns the latter. Our understanding of the molecular interactions behind the RHE is limited. Indeed, it is only in the last decade that it has been confirmed that the key non-covalent interactions are those between the anion of the salt and positively charged groups on the solute. Beyond this, details are sparse: We know little about the magnitude of such interactions and whether they are dominated by Coulombic or dispersion interactions; we know little about the existence of specific ion-pairs that might dominate the precipitation of a solute; and we know little about the mechanisms of the aggregation and precipitation pathway(s). To develop an understanding of these we outline: 1) studies with model hosts designed to probe ion-ion pairing structurally and thermodynamically, and hence reveal details of how these lead to aggregation and precipitation; 2) molecular dynamics (MD) simulations designed to reveal atomistic details of these ion pairs, and the role water plays in modulating their thermodynamics of interaction, and; 3) studies with proteins that, building on our understanding of model hosts and MD simulations, will begin to systematically qualify and quantify how the RHE is manifest in proteins, and the specific ion-ion interactions behind this phenomenon. These studies will address the following scientific questions: What are the specific ion-ion interactions pertinent to the RHE? What are the specific structural features and thermodynamics of these ion-ion interactions? Are there qualitative and quantitative links between the nature of ion pairing and the aggregation and precipitation of small molecules? Do anion-protein interactions influence the structure, stability, and aggregation of proteins in specific, determinable ways? Can the RHE in proteins be used as a signature to characterize/identify proteins? Can the RHE in proteins be attributed to specific anion-protein interactions? Answering these questions will improve our understanding of the solubility of small molecules common to the pharmaceutical industry, and lead to a clearer picture of the often bewildering and contradictory RHE in proteins. This latter point is not only key to determining new ways to purify and crystallize proteins, but is also crucial to understanding the irreversible deposition of proteins in prion diseases and thrombosis.